1. Field of the Invention
The present invention relates to a method for rapidly and easily measuring N-peptide clinically useful for the diagnosis of cardiac incompetence and chronic renal failure, a monoclonal antibody used in the method, and a hybridoma producing the monoclonal antibody.
2. Description of the Related Art
Human atrial natriuretic polypeptide (hANP) is biologically produced as a peptide including 126 amino acids which is referred to as hANP(126) (.gamma.-hANP). This .gamma.-hANP accumulates in intra-atrial granules, and is secreted in response to an atrium pressure stimulus. At this time, .gamma.-hANP is cleaved into hANP(1-98) (N-peptide) and hANP(99-126) (.alpha.-hANP) and simultaneously released into the blood.
The blood level of .alpha.-hANP reflects the cardiac conditions as marker in cardiac incompetence and chronic renal failure, and immunoassay thereof has already been employed for diagnostic purposes. However, the immunoassay has some disadvantages. For example, a very long reaction time (3 days) is required for the assay, and an immunological activity of a-hANP markedly decreases during the process of collecting or storage of a sample due to instability of a-hANP in the blood (See, T. Tsuji et al., Clin. Chim. Acta, 225, 171-177 (1994)). In contrast, N-peptide is more stable and lower in clearance than .alpha.-hANP, the concentration of which is high in the circulatory blood. Therefore, immunoassay of the N-peptide has been developed.
It has already been reported that the immunoassay of the N-peptide which is an N-terminal peptide of the above-mentioned .gamma.-hANP is useful for the diagnosis of cardiac incompetence and chronic renal failure (See, J. A. Sundsfjoed et al., J. Clin. Endocrionol. Metab., 66, 605-610 (1988); H. Itoh et al., J. Clin. Endocrinol., 67(3), 429-437 (1988); M. G. Burckley et al., Clin. Sci. 77, 573-579 (1989); and M. G. Burckley et al., Clin. Chim. Acta, 191, 1-14 (1990)). All of immunoassays for the N-peptide previously reported depend on competitive radioimmunoassay (RIA) methods using one kind of antibody. As a result, the methods require a long period of time (3 to 4 days), a limited temperature condition (about 4.degree. C.), and/or a complicated operation (i.e., Sep-pak C-18 extraction) in which the plasma has to be pretreated (See, J. A. Sundsfjoed et al., supra; and M. G. Burckley et al., (1990) supra). Furthermore, any of these immunoassays have problems such as low precision and poor reproducibility. Thus, the development of a method for rapidly and easily measuring N-peptide has been required.